Chromatographic analysis of oxidized cello-oligomers generated by lytic polysaccharide monooxygenases using dual electrolytic eluent generation

Research on oligosaccharides, including the complicated product mixtures generated by lytic polysaccharide monooxygenases (LPMOs), is growing at a rapid pace. LPMOs are gaining major interest, and the ability to efficiently and accurately separate and quantify their native and oxidized products chromatographically is essential in furthering our understanding of these oxidative enzymes. Here we present a novel set of methods based on dual electrolytic eluent generation, where the conventional sodium acetate/sodium hydroxide (NaOAc/NaOH) eluents in high-performance anion-exchange chromatography (HPAEC) are replaced by electrolytically-generated potassium methane sulfonate/potassium hydroxide (KMSA/KOH). The new methods separate all compounds of interest within 24-45 min and with high sensitivity; limits of detection and quantification were in the range of 0.0 0 01-0.0 032 mM and 0.0 0 02-0.0 096 mM, respectively. In addition, an average of 3.5 times improvement in analytical CV was obtained. This chromatographic platform overcomes drawbacks associated with manual preparation of eluents and offers sim plified operation and rapid method optimization, with increased precision for less abundant LPMO-derived products. (c) 2021 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY license ( http://creativecommons.org/licenses/by/4.0/ )

Automated summary

Research on oligosaccharides, including the product mixtures generated by LPMOs, is growing rapidly. A novel method based on dual electrolytic eluent generation has been developed to separate and quantify these compounds with high sensitivity and improved analytical precision. This new chromatographic platform offers simplified operation and rapid method optimization, overcoming drawbacks associated with manual eluent preparation.